PROJECT SUMMARY Current treatments for HIV-1 infection include suppressive anti-retroviral therapy (ART), which curtails active viral replication to nearly undetectable levels. However, ART fails to cure HIV-1 infection because the virus persists indefinitely in a latent state. While people living with HIV-1 (PLWH) live longer, they are subject to secondary long-term consequences due to the ART regime itself and the presence of integrated proviruses throughout the human genome. Whereas ART facilitated a decrease in the incidence of AIDS defining cancers, an increased incidence of non-AIDS defining cancers (NADC), such as T and B cell lymphomas, has been well documented in the clinics. Despite previous controversies regarding the role of HIV-1 integration site and cancer prevalence, the clinical association between HIV-1 integration in defined sites and NADC prevalence remains largely unexplored. Given these previous findings and gaps in knowledge, the central hypothesis of this proposal is that HIV-1 integration into or nearby select human genes alters their expression thereby causing NADC. Specifically, HIV-1 integration might disrupt the normal expression of HIV-1 Associated GEnes (HAGEs) causing upregulation of proto-oncogenes, downregulation of tumor suppressors and/or altering RNA splicing patterns, consequently rewiring gene expression programs producing abnormal CD4+ T and B cell expansion and/or behaviors leading to NADC. The major goal of this proposal is to interrogate whether select HIV-1 integration sites are associated with increased incidence of NADC in PLWH under suppressive therapy and to predict associated biomarkers. To start accomplishing this goal, we will deploy a multidimensional approach to concurrently profile HIV-1 integration sites and human transcriptomes at the single cell level in a unique cohort available through the AIDS and Cancer Specimen Resource (ACSR). This will be the first ever approach that can simultaneously interrogate the relationship between HIV-1 integration sites and human transcriptomes at the single cell level in participants biospecimens. We already have on hand an impressive molecular and genomic toolset that will be strategic for accomplishing the Specific Aims of this proposal. These include: 1) To deploy a multidimensional single cell approach to simultaneously collect HIV-1 integration sites and human transcriptomes in ACSR participants biospecimens (Aim 1), and 2) to combine the generated datasets to define participants HIV-1 integration-induced biomarkers associated with NADC (Aim 2). Collectively, these studies will reveal viral integration sites over-represented in PLWH with NADC, link individual viral integration sites to altered human gene expression potentially explaining the onset of NADC, and categorize biomarkers linked to select HIV-1 integration events for diagnostic and prognostic outcomes. Future studies using this knowledge will functionally define specific HIV-1 integration ev...